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Other advantages that can be associated with this technique include:
• Low back pressures, due to the low viscosity of the organic-rich mobile phases. This is especially relevant
when using sub-2 μm particles; lower back pressure allows for the use of high flow rates for fast analysis
(a word of warning: the optimum performance may become compromised as it has been observed [13] that
in HILIC higher efficiencies are achieved at lower flow rates than in RPLC. This phenomenon will be illustrated
further on in the guide). Lower back pressure also means that the frictional heating effect – which could be
detrimental when using sub-2 µm particles in RPLC conditions − is not an issue in HILIC, since the pressure is
generally 2−3 times less than in RPLC [15].
• Less peak tailing upon column overloading. This phenomenon is observed when peaks for very polar analytes
(eluted under HILIC conditions) are compared to the peaks that can be obtained for these compounds on silica
columns using highly aqueous conditions (in per aqueous liquid chromatography, PALC). Good retention factors
can be achieved under PALC conditions (by taking advantage of the hydrophobic character of siloxane groups at
the surface of the silica), however the underlying adsorption mechanism is heterogeneous and involves active
adsorption sites; this results in column overloading being experienced when analyzing strongly retained
samples (k > 2) [16].
• Compatibility with SPE extracts, if in 100% aprotic organic solvent. These can be directly injected
without the need to dry down and reconstitute in mobile phase. Since the organic solvents are weak eluents
under HILIC conditions, polar analytes are accumulated on a narrow zone near the column inlet (phenomenon
described as ‘sample on-column’ focusing) [17].
